Analysis of glucose metabolism in farmed European sea bass (Dicentrarchus labrax L.) using deuterated water

Effects of food-deprivation and refeeding on the regulation and sources of blood glucose appearance in European seabass (Dicentrarchus labrax L.)

“…The supernatant was desalted by passage through sequential columns containing Dowex® 50WX8 (hydrogen form, Sigma-Aldrich) and Amberlite® IRA-67 (free base, Fluka, Sigma-Aldrich), then lyophilized and stored at −20°C. In order to optimize the signal resolution in the NMR spectra, glucose was converted into its derivative, monoacetone glucose (MAG), as previously described (Viegas et al, 2011). Hepatic glycogen was extracted by precipitation with ethanol after alkaline tissue hydrolysis and glycogen was broken into its glycosyl units with amyloglucosidase from Aspergillus niger (glucose-free preparation, Sigma-Aldrich) as previously described (Viegas et al, 2012).…”

“…Proton-decoupled 2 H NMR spectra of MAG samples were obtained at 50°C with a 14.1 T Varian spectrometer (Varian, Palo Alto, CA, USA) equipped with a 3 mm broadband probe with the observe coil tuned to 2 H as previously described (Viegas et al, 2011). Approximately 12,000 to 17,000 scans were acquired per sample (corresponding to 6-8 h of collection time) and field-frequency drift was compensated by the Scout Scan software (Varian).…”

“…Yet, these by-difference calculations as defined by the equations presented in this work rely on the assumption that G6P fully exchanges with body water such that the enrichment of hepatic G6P position 2 is equivalent to that of body water (Viegas et al, 2011). To the extent that this exchange is incomplete, the fractional synthetic rate of G6P from non-gluconeogenic precursors is underestimated.…”

“…For example, in seabass the activities of hepatic gluconeogenic enzymes were not suppressed by chronic high-CHO feeding thereby contributing to postprandial hyperglycemia (Enes et al, 2006(Enes et al, , 2008b. Meanwhile, the transition from feeding to fasting is characterized by a steep drop in plasma glucose concentrations (Gutiérrez et al, 1991;Echevarría et al, 1997;Pérez-Jiménez et al, 2007;Viegas et al, 2011;Pérez-Jiménez et al, 2012) among other tissue-dependent effects (Blasco et al, 2001). Other fish species can tolerate extended periods of starvation without apparently suffering from hypoglycemia like Brycon cephalus (Figueiredo-Garutti et al, 2002), gilthead seabream Sparus aurata (Sangiao-Alvarellos et al, 2005;Polakof et al, 2006) or Senegalese sole Solea senegalensis (Costas et al, 2011).…”

“…1). This approach was recently applied in seabass (Viegas et al, 2011) revealing that gluconeogenesis was the main source of hepatic G6P production regardless of nutritional state. However, for fasted fish, where blood glucose appearance was expected to be completely dependent on G6P hydrolysis, the fraction of blood glucose derived from G6P was unexpectedly low, accounting for only~70% of the total.…”